PROJECT SUMMARY ALI/ARDS is a serious condition with high mortality rates, and more complete understanding of pathologic mechanisms mediating lung vascular inflammation and barrier dysfunction in ARDS is critical for development of efficient therapeutic approaches to confront this devastating disease. Previous studies by our and other groups revealed pronounced anti-inflammatory and barrier enhancing effects of cyclic AMP elevating agonists on pulmonary vascular endothelium, which accelerated ALI recovery. Endogenous cAMP levels also appear to be critical for the maintenance of endothelial cell anti-inflammatory status and barrier function. However, septic and ALI conditions lead to impairment of cAMP homeostasis, which may contribute to severity of endothelial dysfunction and lung injury in ARDS. The importance of this mechanism is supported by beneficial effects of pharmacological inhibition of cAMP hydrolyzing enzyme, phosphodiesterase (PDE) in preclinical models of septic ALI. Despite these encouraging results, precise molecular mechanisms of PDE activation in inflammatory conditions still remain to be elucidated. Coagulation and inflammation are activated by the same types of challenges and correlate both temporally and spatially in different pathologies, but mechanistic interactions between these two processes are incompletely understood. Fibrinogen is a key component of the coagulation system. Increased levels of fibrinogen and fibrin deposition are distinctive features of advanced ALI and septic syndromes. Fibrinogen directly interacts with ?5?1 integrin adhesion receptor expressed by pulmonary endothelial cells. Our exciting pilot studies show that this interaction may augment EC inflammation and barrier dysfunction caused by bacterial pathogens via recruitment of PDE4 to the ?5-integrin associated signaling protein complex. This proposal will investigate for the first time the molecular mechanism of synergy between ARDS-relevant coagulation component fibrinogen and lung EC inflammation caused by bacterial particles. Aim-1 will employ in vitro and in vivo models of ALI caused by Gram-positive bacterial particles to evaluate fibrinogen role in lung vascular endothelial dysfunction and severity of lung injury. Aim-2 will investigate assembly and activation of ?5-integrin-ILK-paxillin signalosome and evaluate its role in the mediation of fibrinogen-induced exacerbation of endothelial dysfunction and lung inflammation. Aim-3 will study targeting of PDE4 to fibrinogen-activated ?5-integrin-ILK-paxillin signalosome and its significance for PDE4- dependent suppression of intracellular cAMP and augmentation of HKSA-induced ALI.